PB2062: PAROXYSMAL NOCTURNAL HEMOGLOBINURIA SCREENING AFTER INTRODUCTION OF HIGH-SENSITIVITY DIAGNOSTICS – A SINGLE-CENTER 10-YEAR EXPERIENCE
Authors: Eliza Głodkowska-Mrówka, Justyna Spychalska, Magda Duńska, Anna Myslinska, Monika Majewska-Wierzbicka, Edyta Klimczak-Jajor, Malgorzata Uhrynowska
Journal: HemaSphere (2023)

Topic: 12. Bone marrow failure syndromes incl. PNH - Clinical Background: Paroxysmal nocturnal hemoglobinuria (PNH) is a rare clonal hematopoietic stem cell disorder characterized by lack or low expression of the cell-surface glycophosphatidylinositol (GPI) and GPI-anchored proteins. The high-sensitivity multiparameter flow cytometry is recommended to detection GPI-deficient cells and the screening of PNH especially in patients with bone marrow failure syndrome: hypoplastic or aplastic anemia (AA), myelodysplastic syndrome (MDS) and other cytopenias of unknown etiology. FLAER-based tests on leukocytes and 2-color tests on erythrocytes allow to detect PNH phenotype with 0.01% sensitivity. Aims: Retrospective evaluation of high-sensitivity PNH testing at the single hematology center over 10 years. Methods: Data from 2790 patients first time examined for PNH at the Institute of Hematology and Transfusion Medicine in Warsaw, Poland from 01.01.2013 until 31.12.2022 were included to this retrospective analysis. Patients’ peripheral blood samples (EDTA) were tested within 48 h after collecting. Leukocytes were analyzed using 2 tubes method using 4-color cocktail: FLAER/CD24/CD45/CD15 for neutrophils and FLAER/CD14/CD45/CD34 (2013-04.2016) or FLAER/CD14/CD45/CD64 (05.2016-2022) for monocytes. Erythrocytes were analyzed with 2-color cocktail CD235a/CD59. Samples acquisition and analysis were performed on a BD FACSCalibur™ cytometer with BD CellQuest Pro™ software. Acquisition for each test was up to 250 000 cells. PNH clone was expressed as a sum total of type II cells (partial GPI deficiency) and type III cells (total loss of GPI expression, GPI-). Detailed analysis was performed in a cohort of 346 (12.4% of all tested) patients PNH+ confirmed with high-sensitivity assay. A case was defined to be PNH+ when GPI- were found in ≥2 lineages of cells (neutrophils, monocytes and/or erythrocytes) at a level of ≥0.02%. Results: Over the studied decade, on average, 35 new PNH cases were detected a year with 15 new cases/year for PNH clone size ≤1%. In PNH+ group, neutrophils PNH clone size varied from 0.02 to 99.80%. In 153 (44%) patients PNH clone was ≤1%. Large PNH clone (>50%) was observed in 48 cases. Of these, there were 3/135 AA (2% of total AA PNH+), 10/81 other cytopenias (12%), 9/38 MDS (24%), and 19/25 hemolysis (76% of total hemolytic anemia PNH+). Screening was performed due to AA (39%), cytopenias of unknown etiology (23%), hypoplasia (15%), MDS (11%), hemolysis (7%); thrombosis (1%) and no data/other (4%). Predominant initial diagnosis was cytopenia (n=268; 77%), including AA (n=135; 50.4%), hypoplastic anemia (n=52; 19.4%) and other cytopenias of unknown etiology (n=81; 30.2%). Less frequent diagnoses included MDS (n=38; 11%), hemolytic anemia (n=25; 7%), and thrombosis (n=2; 1%). Summary/Conclusion: This is the first evaluation of 10-year experience in PNH screening with high-sensitivity detection assay in a Polish cohort. Our findings confirm the need to test high-risk groups of patients (especially with bone marrow failures) with high-sensitivity PNH tests as recommended by ICCS guidelines and provide valuable insight into clinical characteristics of PNH patients at diagnosis. Noteworthy most hemolytic anemia patients had large PNH clones (>50%) that highlight the need to include PNH screening in this population at earlier stages of diagnostic workup to shorten the time to final diagnosis. Keywords: Flow cytometry, Paroxysmal nocturnal hemoglobinuria (PNH), Screening, Diagnosis